The major objective of the proposed research is to establish the pattern of cyclic nucleotide metabolism in mouse and rat Sertoli cell cultures. The cellular content of cyclic nucleotides (cAMP and cGMP) and adenylate and guanylate cyclases and the accumulation of cyclic nucleotides in Sertoli cell culture media will be determined by radioimmunoassay methods before and after treatment with FSH and DHT both alone and indifferent temporal combinations. The distribution in Sertoli cell monolayers of cAMP, cGMP and both catalytic and regulatory subunits of cAMP-dependent protein kinases will be determined by immunocytocemistry before and after hormonal treatment. It is hoped that these experiment will determine (1) the specific cell distribution sites of cAMP, cGMP and protein kinase subunits in control Sertoli cells and (2) the distinctive effect of FSH and DHT on the distribution patterns of cyclic nucleotides and protein kinases. In addition, structural aspects of RNA synthesis will be investigated in Sertoli cell cultures by electron microscope autoradiography before and after FSH and DHT treatment. Tritiated uridine will be used to investigate: (1) the time course labeling of hormone-induced gene activity, (2) the distribution sites of newly synthesized RNA species and (3) the relationship between RNA synthesis and cyclic nucleotide metabolism. Finally, we will detemine the immunocytochemical distribution of cyclic nucleotides and protein kinase subunits in prepuberal testes interacting in vitro with Sertoli cell cultures for variable periods of time. This approach intends to determine the possibility that the hormone-dependent Sertoli cell gene expression may trigger a precocious initiation of meiosis in prepuberal testis or affect meiosis in puberal testis. All of the above studies should contribute to a greater understanding of the hormone-cyclic nucleotide interaction in the differentiating Sertoli cell population as a whole. The results may become relevant for the study of the control mechanism underlying the spermatogenic cell differentiation process.